FLAVIN ADENINE-DINUCLEOTIDE AND FLAVIN MONONUCLEOTIDE METABOLISM IN RAT-LIVER - THE OCCURRENCE OF FAD PYROPHOSPHATASE AND FMN PHOSPHOHYDROLASE IN ISOLATED-MITOCHONDRIA

In order to gain some insight into mitochondrial flavin biochemistry, rat liver mitochondria essentially free of lysosomal and microsomal co ntamination were prepared and their capability to metabolise externall y added and endogenous FAD and FMN tested both spectroscopically and v ia HPLC. The existence of two novel mitochondrial enzymes, namely FAD pyrophosphatase (EC 3.6.1.18) and FMN phosphohydrolase (EC 3.1.3.2), w hich catalyse FAD-->FMN and FMN-->riboflavin conversion, respectively, is shown. They differ from each other and from extramitochondrial enz ymes, as judged by their pH profile and inhibitor sensitivity, and can be separated in a partial FAD pyrophosphatase purification. Digitonin titration and subfractionation experiments show that FAD pyrophosphat ase is located in the outer mitochondrial membrane and FMN phosphohydr olase in the intermembrane space. Since these enzymes can metabolise e ndogenous FAD and FMN, which are made available by using both Triton X -100 and the effector oxaloacetate, a proposal is made that FAD pyroph osphatase and FMN phosphohydrolase play a major role in mitochondrial flavoprotein turnover.